Selection and Clonal Propagation of High Artemisinin Genotypes of Artemisia annua

Abstract

Artemisinin, produced in the glandular trichomes of Artemisia annua L. is a vital antimalarial drug effective against Plasmodium falciparum resistant to quinine-derived medicines. Although work has progressed on the semi-synthetic production of artemisinin, field production of A. annua remains the principal commercial source of the compound. Crop production of artemisia must be increased to meet the growing worldwide demand for artemisinin combination therapies (ACTs) to treat malaria. Grower artemisinin yields rely on plants generated from seeds from open-pollinated parents. Although selection has considerably increased plant artemisinin concentration in the past 15 years, seed-generated plants have highly variable artemisinin content that lowers artemisinin yield per hectare. Breeding efforts to produce improved F1 hybrids have been hampered by the inability to produce inbred lines due to self-incompatibility. An approach combining conventional hybridization and selection with clonal propagation of superior genotypes is proposed as a means to enhance crop yield and artemisinin production. Typical seed-propagated artemisia plants produce less than 1% (dry weight) artemisinin with yields below 25 kg/ha. Genotypes were identified producing high artemisinin levels of over 2% and possessing improved agronomic characteristics such as high leaf area and shoot biomass production. Field studies of clonally-propagated high-artemisinin plants verified enhanced plant uniformity and an estimated gross primary productivity of up to 70 kg/ha artemisinin, with a crop density of one plant m-2. Tissue culture and cutting protocols for the mass clonal propagation of A. annua were developed for shoot regeneration, rooting, acclimatization, and field cultivation. Proof of concept studies showed that both tissue culture-regenerated plants and rooted cutting performed better than plants derived from seed in terms of uniformity, yield, and consistently high artemisinin content. Use of this technology to produce plants with homogeneously-high artemisinin can help farmers markedly increase the artemisinin yield per cultivated area. This would lead to increased profit to farmers and decreased prices of ACT.

Seasonal and Differential Sesquiterpene Accumulation in Artemisia annua Suggest Selection Based on Both Artemisinin and Dihydroartemisinic Acid may Increase Artemisinin in planta

“…Plants that have higher DHAA than ART (Chinese) may be crossed with plants that have higher ART than DHAA (Brazilian) to generate plants with even higher ART than either parent, as these genotypes may have an ideal combination of enzymatic systems needed to transform DHAA into ART. Although the cross could not be genetically confirmed, but based on their low selfing rate, crosses between Brazilian and Chinese plants, done by pairing plants synchronized to flower in Indiana (Purdue), and selected in Georgia were recently reported to produce 2.16% ART (Wetzstein et al, 2018 ). Alternatively, genotypes with high DHAA may be genetically engineered to overexpress enzymes needed to convert DHAA into ART, if eventually demonstrated that indeed enzymes are involved in the end of the ART pathway in vivo (Zhu et al, 2014 ).…”

Using Micropropagation to Develop Medicinal Plants into Crops

“…Selection of genotypes with high artemisinin co wild populations resulted in lines containing up to 1.4 percent artemisinin basis). The leading commercial source, 'Artemis,' exhibited extensive var bolic and agronomic traits; artemisinin content on a µg/mg dry basis for in ranged 22-fold, plant fresh weight varied 28-fold, and leaf area ranged 9-f While Ferreira and Janick [32] found that the in vitro production of a never be commercially feasible, Wetzetein et al [33] suggested that cultiv propagated high-yielding artemisinin plants with levels above 2% and i nomic traits (high leaf area and shoot biomass production) may reach pro kg/ha artemisinin using a crop density of 1 plant m −2 . We include in Table pharmaceutical plant species classified as small therapeutic molecules S their micropropagation protocols to produce elite clones for higher yield other success story.…”

“…While Ferreira and Janick [32] found that the in vitro production of artemisinin will never be commercially feasible, Wetzetein et al [33] suggested that cultivation of micropropagated high-yielding artemisinin plants with levels above 2% and improved agronomic traits (high leaf area and shoot biomass production) may reach productivity of 70 kg/ha artemisinin using a crop density of 1 plant m −2 . We include in Table 1 examples of pharmaceutical plant species classified as small therapeutic molecules STM's (18) and their micropropagation protocols to produce elite clones for higher yields.…”

Chloroplast genome transformation of medicinal plant Artemisia annua

“…These plantlets after analysing for transgenes integration by PCR, Southern blot and eGFP protein expression by Western blot analysis were transferred to greenhouse. The transplastomic plants were further clonally propagated as reported (Wetzstein et al , 2018). Transplastomic plants thrive well in the greenhouse, and growth was comparable to WT plants (Fig.…”